Medulloblastoma (MB) is the most common malignant brain tumor in children. Recent studies have divided MB into four molecular subgroups: WNT, SHH, Group 3, and Group 4. While WNT and SHH tumors have a relatively favorable prognosis, Group 3 tumors (characterized by overexpression of the MYC oncogene) are much more aggressive and almost invariably fatal. The first mouse model of Group 3 MB was recently established in our lab by infecting neural stem cells (NSCs) with viruses encoding Myc and a dominant- negative form of the p53 tumor suppressor (DNp53), and then transplanting these cells back into the cerebellum of immunodeficient mice. Although the resulting tumors resemble human Group 3 MB, this model does not precisely recapitulate the genetics of the human disease, as most human Group 3 tumors do not exhibit mutation or loss of p53. Thus, a major goal of our research has been to identify second hits that can cooperate with MYC and are relevant to human MB. Recent whole genome sequencing analysis has identified two chromosomal loci that are hotspots for rearrangement in Group 3 MB. Rearrangements at these loci activate the zinc-finger transcriptional repressors Gfi1 and Gfi1b. Importantly, our preliminary data show that both Gfi1 and Gfi1b can cooperate with Myc to drive MB formation in mice. Based on the striking prevalence of these events in human Group 3 MB as well as previous reports that Gfi1 cooperates with Myc in leukemia, we hypothesize that Gfi proteins are key regulators of tumorigenesis and important therapeutic targets in Group 3 MB. To address this hypothesis, we propose to (1) Identify the cofactors and transcriptional targets that mediate the tumor-promoting effects of Gfi1/1b, (2) Determine if Gfi1 and Gfi1b are required for tumor maintenance, and (3) Identify therapeutic agents that inhibit the growth of Gfi1/1b-activated tumors. The proposed studies will provide insight into the molecular mechanisms of tumorigenesis in new, genetically- relevant models of Group 3 MB and help identify novel targets for therapy of this disease.